The work performed in this proposal will help in the development of tissue engineered cartilage constructs, to be commercialized as the TEC-AriTM method. The product is intended to be a tissue engineering service available to health care providers that involves the use of autologus cells to construct in vitro articular cartilage. The neotissue, produced using a self-assembling process, can be subsequently used to treat chondral, focal lesions. According to the CDC, more than 150,000 surgical procedures are preformed annually to treat knee lesions. The current state of the art treatment involves delivery of a cell suspension, which is not biomechanically functional, to the defect site. The self-assembling process is a novel tissue engineering method shown to be capable of producing neotissue without the use of an exogenous scaffold. Preliminary studies have shown that the mechanical properties of chondrocyte self-assembled neotissue can reach values on par with native adult tissue within 4 weeks. The objective of this application is to evaluate the feasibility of forming tissue engineered articular cartilage constructs using stromal mesenchymal stem cells (MSCs), in a self-assembling process. The study's hypotheses are 1) the self-assembling process, when applied to MSCs will result in tissue engineered articular cartilage constructs of clinically relevant dimensions (discs more than 1 mm thick and 5 mm in diameter); and 2) the functional properties of the self-assembled MSC constructs will be tantamount to those of native articular cartilage. To examine these hypotheses, the following specific aims have been proposed: 1) to form tissue engineered constructs using MSCs and the self-assembling process; and 2) to compare the functionality of self-assembled MSC constructs to self-assembled chondrocyte constructs and to native tissue. MSCs, differentiated into chondrocytes, will be allowed to self-assemble to form tissue engineered constructs. These constructs will then be cultured with or without confinement and with or without sequential seeding to determine an optimal construct formation method that will be used for the second specific aim. 'Optimal' is defined as the set of conditions producing properties closest to native tissue in terms of extracellular matrix composition and biomechanical properties. An optimal method must also yield constructs 1 mm thick or thicker. Self-assembled constructs formed using MSCs and native chondrocytes using the optimal method determined in aim 1 will then be cultured and compared to native articular cartilage in terms of biochemistry and biomechanics. Completion of this work is pivotal toward commercialization of the TEC-Ari method which is expected to address the yet unsolved clinical problem of articular cartilage defects. PUBLIC HEALTH RELEVANCE: The work performed in this proposal will help foster new strategies for articular cartilage regeneration. Advancing our clinical capabilities to treat cartilage defects, by engineering articular cartilage, which is then integrated functionally into the joint, will significantly improve the quality of life of millions of people who suffer with cartilage ailments. Ariston Medical's innovative approach to cartilage regeneration using a self- assembling process has the potential to reduce the burdens and disability caused by cartilage degeneration. [unreadable] [unreadable] [unreadable]